The application of nanotechnology to cancer research is an exciting frontier in the efforts to develop novel approaches for cancer detection and treatment. Although the feasibility of using nanoparticles for cancer detection and drug delivery has been demonstrated in several laboratories [1-3], a major obstacle limiting its clinical application is that non-tumor targeted nanoparticles are unable to reach sufficient concentration in the tumor site to either produce a strong signal for tumor imaging or to carry optimal amounts of therapeutic agents into tumor cells.
Breast cancer is the most common type of cancer and a leading cause of death among women. Crucial factors that would increase patient survival are early detection and effective treatment. The development of novel approaches for detecting and treating breast cancer are urgently needed to increase patient survival. Furthermore, since cancer metastasis is the main cause for the mortality in breast cancer patients. Any new approaches for detection and targeted treatment of distant metastases should also significantly improve patient prognosis.
Although various imaging technologies and modalities have been widely used for management of cancer including diagnosis and treatment monitoring, conventional tumor imaging methods such as magnetic resonance imaging (MRI), X-ray computerized tomography (CT) or even positron emission tomography (PET) have their limitations in both specificity and sensitivity of cancer detection [4-6]. Increasing evidence suggests that the use of targeted imaging probes enhances signal intensity in the tumor, increasing the sensitivity of the detection [7-10]. Furthermore, imaging agents that target changes in the tumor environment, such as tumor endothelial cells and intra-tumor stromal cells, should further augment tumor imaging signals [11].
It is clear that selective delivery of therapeutic agents into a tumor mass has the potential to minimize toxicity to normal tissues, while improving bioavailability of cytotoxic agents in the tumor [12, 13]. Antibodies, ligands and peptides that target to cell surface molecules, which are highly expressed in tumor cells or tumor endothelial cells have been used to deliver therapeutic agents, showing promise in achieving tumor specific cytotoxicity [3, 14]. An important way to improve the delivery of therapeutic agents is to limit the size of the delivery complex in many currently used delivery systems such as antibody-conjugates, liposomes and other macromolecules, since it is well known that solid tumors will show very poor bio-distribution of the large molecules due to the dysfunctional blood and lymphatic vessels and compressive pressure in the tumor [15-17]. Therefore, the use of drug delivery vehicles with sizes of a few nanometers will enhance the efficiency of delivery of therapeutic agents into solid tumors.
Additionally, tumor imaging plays a key role in helping clinicians to detect solid tumors, to determine tumor recurrence and to evaluate the response of the tumors to therapeutic reagent. The combination of imaging technology and tumor biology has created a “molecular imaging” field with new applications in all imaging modalities. The methods for high-resolution in vivo imaging using mainly three types of imaging probes: radio-labeled, magnetic and optical probes for PET and single photon emission tomography (SPECT); MRI and spectroscopy; and optical imaging techniques, including fluorescence-mediated tomography (FMT) and near-infrared fluorescence (NIRF) reflectance imaging. Although different modalities vary in imaging sensitivity and resolution, the technical challenge in improving target specificity and sensitivity is common. In a clinical practice, for example, 18F-fluoro-2-deoxy-D-glucose (FDG) and Gd(III)-aminobenzyl-diethylenetriaminepentaacetic acid (Gd-DTPA) contrast agent are used commonly for PET and MRI, respectively. However, both have significant limitations in sensitivity and specificity in delineating tumor and detecting cancer cells in the early stage of development of tumor [5, 44, 45]. Recently, tumor-targeted optical, radio- or magnetic probes have been generated and the feasibility of those imaging probes was examined in both animal tumor models and in clinical studies [1, 7, 10, 46, 47]. Those results show that tumor-targeted imaging probes can increase the localization of the image probes in tumors while reducing their uptake in normal tissues. However, to develop a promising tumor imaging approach to clinical applications, several important issues have to be addressed in the research laboratory. The most important issues include: 1) developing of imaging probes that emit a strong signal to improve sensitivity of detection; 2) targeting probes to cellular receptors that are highly expressed in human tumor cells or tumor environments and demonstrating that there is low toxicity to normal organs and tissues; and 3) developing an effective delivery system to direct the imaging probe to the targeted tumor or cancer cells.
At present, three types of imaging probes are used for in vivo imaging: optical, magnetic and radio labeled probes. Optical image probes use organic fluorescence dyes, fluorescence proteins, and semiconductor quantum dots. Emerging as a new class of fluorescent probes for in vivo biomolecular and cellular imaging, these quantum dots (QDs) are tiny, nanometer-scale light-emitting particles. In comparison with organic dyes and fluorescent proteins, quantum dots have unique optical and electronic properties such as size-tunable light emission, improved signal brightness, resistance against photobleaching, and ability to simultaneous excite multiple fluorescence colors [48]. These properties are most promising for improving the sensitivity of molecular imaging and quantitative cellular analysis by 1-2 orders of magnitude. Nie et al. first reported that it is feasible to simultaneously target and image prostate tumors in living animal models using bioconjugated prostate membrane antigen-targeted QDs [1]. This new class of QD conjugates contains an amphiphilic triblock copolymer layer for in vivo protection and multiple PEG molecules for improved biocompatibility and circulation, making it highly stable and able to produce bright signals. This triblock copolymer layer is designed so that it can have multiple active functional groups for conjugation of different tumor targeting moieties and therapeutic agents on the same nanoparticle. Another advantage is that multicolor QD probes can be used to image and track multiple tumor markers simultaneous, which will most likely increase the specificity and sensitivity of cancer detection.
Recently, QDs producing near infrared (NIR) signals have also been developed [49, 50]. NIR light penetrates much more deeply into tissues, compared to visible fluorescence, and allows detection of signals inside animals. The feasibility of detection of NIR signals in animal tumor models has been demonstrated using fluorescent dye Cy 5.5-labeled RGD peptide or an enzyme-activated Cy 5.5 NIR signal [10, 51, 52]. Detection of QD NIR signals in sentinel lymph node in large animals real time has also been demonstrated [50, 53]. A major advantage of NIR QDs is that emissions of those QDs are well beyond the spectral range of autofluorescence in tissues, thus resulting in imaging with a high signal: background ratio [53].
In comparison to optical imaging, magnetic resonance imaging (MRI) has lower sensitivity when applied for molecular and cellular imaging. However, it has super imaging resolution and deep tissue penetration for visualizing abnormalities in small animal and human using tissue water molecules as signal sources. It is a non-invasive imaging modality and is routinely used in the clinic for diagnostic imaging. To obtain contrast enhancement and signal amplification, paramagnetic contrast agents are often used. Although Gd-DTPA, a blood-pool contrast agent, is widely accepted in the clinical uses, superparamagentic iron oxide (SPIO or IO) nanoparticles are emerging as a new generation of MRI contrast agent for the development of target specific contrast agent, because it has a long blood retention time, low toxicity and biodegradability. The IO nanoparticles possess unique paramagnetic properties, which generate significant susceptibility changes resulting in strong T2 and T*2 contrast [45, 54]. In addition, the surface coating molecules used for the IO nanoparticles can be conjugated to the biomolecule to provide target specific interaction to the cell [54]. Several recent studies have demonstrated that the IO nanoparticles can be internalized by various cell lines including cancer cells to allow magnetically labeling of the targeted cell. When internalized by cells, the IO nanoparticles are able to generate MRI contrast that enables single-cell MR detection [55]. At present, non-targeted IO particles has been used in clinic and is proven to be safe for human use.
Over the past years, significant efforts have gone toward developing a target specific MRI contrast agent based on the formulation of the IO nanoparticles [56-59]. However, several obstacles remain to be overcome. The major challenge is to develop a surface coating material that not only can stabilize the nanoparticles but also to provide active functional groups available for controllable bioconjugation of “probe” ligands. Traditional ligands (e.g., dextran) that are used for the stabilization of magnetic nanocrystals often have weak ligand-particle interactions, so they can be easily detached from the nanocrystal surface, leading to nanoparticle aggregation and eventually precipitation even under physiological conditions or even just during storage. Since further derivatization is needed, such a weak interaction between ligand and particle may not withstand the required reaction conditions.
Therefore, a heretofore-unaddressed need exists in the art to address the aforementioned deficiencies and inadequacies.